Methods for processing cell block

ABSTRACT

A system for making cell blocks includes a cell block cassette and processing station, the cassette including a main body having a having a collection aperture formed therein and a filter assembly removably attached to the main body, the filter assembly defining a collection well in communication with the collection aperture, and having a filter positioned across a bottom surface of the collection well, the filter configured to retain cellular matter carried in a fluid that is dispensed into the collection well and flows across the filter. The cell block processor has a cassette interface removably seating the cell block cassette, and a sensor positioned or positionable to detect and monitor a fluid level in the collection well. The processing station includes an automated fluid delivery system operable to dispense a fluid into the collection well, and a controller operatively coupled to the fluid delivery system, wherein the controller causes the fluid delivery system to selectively dispense fluids into the collection well based at least in part on a flow rate across the filter determined at least in part based on changes in the monitored fluid level in the collection well.

RELATED APPLICATION DATA

The present application claims the benefit under 35 U.S.C. § 119 to U.S.Provisional Patent Applications Ser. Nos. 60/822,449, filed on Aug. 15,2006, and 60/863,941, filed on Nov. 1, 2006. The foregoing provisionalapplications are each incorporated by reference into the presentapplication in their entirety.

FIELD OF THE INVENTION

The present inventions pertain to systems and methods for preparingcells for microscopic examination, and more particularly to automatedand semi-automated systems and methods for embedding cellular materialsand tissue fragments within a paraffin substrate that may be thereafterthinly-cut using a standard microtome, for microscope examination.

BACKGROUND

It is useful for diagnosing or detecting a disease process to perform ahistologic or cytologic examination of a tissue cell sample using alight microscope. This requires that a tissue (cellular material) samplemust first be retrieved from the patient, and then processed formicroscopic examination. A number of minimally invasive techniques areavailable for retrieving and collecting cell samples from a patient,e.g., by using a fine needle aspiration biopsy, or by brushing bodycavity surfaces accessible through minimally invasive endoscopictechniques. A variety of cell sample processing techniques are alsoknown, such as the Cytospin® technique and the Thin-Prep® technique, fordepositing cellular materials and tissue fragments directly onto amicroscope slide. Another technique, commonly referred to as a cellblock preparation, immobilizes cellular materials and/or small tissuefragments within a solid support structure, typically paraffin. Thinsections of the cell block are then cut with a microtome and mountedonto a microscope slide for examination.

U.S. Pat. No. 6,913,921 discloses and describes methods and apparatusfor cell block preparation, including providing a tissue collectioncassette that serves a dual function of capturing cellular sample matterand providing a fluid pathway through which the cell processing andembedding reagents can flow. The cellular sample material is provided inan aqueous solution or a liquid cell preservative, which is passedthrough the tissue cassette across a filter that traps the cells andtissue fragments. A reagent flow pathway is configured to sequentiallypass embedding reagents (alcohol, xylene, eosin stain) and liquefiedparaffin through the tissue cassette and the cell sample alreadydeposited on the filter. Once the paraffin is cooled, the filter ispeeled away, leaving a paraffin “disk” protruding from the tissuecassette, with embedded cellular matter positioned at the end of thedisk in a plane at which a tissue section can be cut using a standardmicrotome for microscope examination.

While representing an improvement over the then-state of the art forcell block preparation, the methods and apparatus disclosed in the '921patent remain labor intensive, requiring manual operation andsupervision, in particular, for determining when a sufficient quantityof cellular material has been gathered on the filter. Further, the cellsample fluid “pathway” (tubing and sample port) must be replacedfollowing each use to avoid contamination of subsequent samples, and thefilter-to-cassette connection relies on a relatively thick o-ring inorder to create a sufficient length of paraffin cell block for latermicrotome slicing.

SUMMARY OF THE DISCLOSED INVENTIONS

Systems and methods are disclosed herein for the efficient creation ofparaffin-embedded cell blocks, including several improvements over themethods and the apparatus disclosed in U.S. Pat. No. 6,913,921, such as(but not limited to) substantially automated cell block creation thatdoes not require human oversight, an innovative two-piece cassette andfilter assembly, more consistent cellular matter quantities in thecreated cell blocks, shorter processing time, reduced use of hazardousreagents, and more fully encapsulated cell blocks to preserve nucleicacid integrity.

In an exemplary embodiment, a cell block preparation system includes atwo-piece cell block cassette, a cell block processing station and afinishing station. The cell block cassette includes a main cassette bodyhaving a collection aperture formed therein, and a filter assemblyremovably attached to the main cassette body and defining a samplecollection well in communication with the collection aperture. A filterpositioned across a bottom surface of the collection well is configuredto retain cellular matter (e.g., cervical cells) carried in a samplefluid (e.g., preservative solution) that is dispensed into thecollection well and flows across the filter. In embodiments disclosedherein, the filter assembly comprises a single-piece housing having abase portion forming the sealing surface, and a neck portion extendingfrom the base portion. The neck portion of the filter assembly housingdefines a perimeter of the collection well and has a top end that makesan interference fit with an annular grove located in a bottom surface ofthe main cassette body in order to detachably attach the filter assemblyto the main cassette body. The filter assembly housing further comprisesa filter support retention portion extending from an underside of thebase portion, with a thermally conductive porous filter support member(e.g., a sintered metal disc or a metal screen) retained therein, thefilter being positioned atop the filter support member within thecollection well. The base portion sealing surface slopes at a downwardangle from the neck portion, such that an outermost edge of the sealingsurface extends to (or beyond) an outermost edge of the filter supportretention portion in order to form a seal with the waste chamberinterface (described below) on the processing station.

The processing station has a cassette interface that removably seats thecell block cassette, with a sensor (e.g., an ultrasound sensor)positioned or positionable to detect a fluid level in the samplecollection well from which the flow rate across the filter (which isrelated to the amount of cellular material retained on the filtersurface) may be calculated. A waste chamber compartment underlies thecassette interface, the filter assembly sealing surface sealablyengaging waste chamber interface when the cell block cassette is seated(and latched) in the cassette interface. Respective pressurized air andvacuum sources may be selectively placed in communication with theinterior of the waste chamber, so as to force air back through thefilter and into the collection well; or alternatively to draw air,fluid, or both, from collection well, through the filter, and into thewaste chamber interior. The sample fluid is drawn across the filter withthe assistance of the vacuum, which may be interrupted for administeringback air pressure pulses to momentarily push the cellular material awayfrom the filter surface and allow for fluid in the collection well todrain.

The processing station includes an automated fluid delivery system and acontroller that causes the fluid delivery system to dispense samplefluid from a sample vial into the cell block cassette collection well.In one embodiment, the automated fluid delivery system comprises anautomated arm assembly including a pipette tip holder configured toselectively retrieve, carry and dispose of pipette tips. A suctionsource is coupled to the pipette tip holder, such that an open proximalend of a pipette tip held by the pipette tip holder may be selectivelyconnected to the suction source for aspirating sample fluid, andliquefied paraffin. The processing station is equipped with empty(sterile) pipette tips, a sample vial interface for holding a samplevial containing cellular material suspended in a liquid carrier, asupply of liquefied paraffin (e.g., a heated paraffin bath), and liquidreagent sources (e.g., xylene and isopropyl alcohol), which are eachalso connected to the pipette holder for being dispensed through apipette tip into the sample collection well.

For cell block processing, the automated arm assembly is configured toselectively retrieve a pipette tip, position the retrieved pipette tipto aspirate fluid from a sample vial seated in the sample vialinterface, and then dispense the drawn sample fluid into the samplecollection well. The controller monitors a fluid level in the collectionwell as a function of time and, based on the fixed dimensions of thecollection well, the controller calculates a flow rate across the filterbased on the fluid level output signals received from the sensor. Thesensor is preferably fixed relative to the cassette interface, with theautomated arm assembly being movable relative to the sensor. In the caseof using an ultrasonic sensor, the sensor should be directedorthogonally to the fluid surface in the collection well for accuratetime of flight reflection readings. Depending on the calculated fluidflow rate across the filter, the controller causes the automated armassembly to continue to draw and dispense sample fluid from the samplevial into the collection well, respectively, until the flow rate (evenwith occasional back air pressure “burping” through the filter) is suchthat the controller determines a desired threshold amount of cellularmaterial has been deposited on the filter. Notably, the cell blocks mayadditionally include larger tissue fragments manually placed on thefilter and then augmented by aspiration of additional cellular materialfrom the sample fluid.

Once the controller determines that desired threshold amount of cellularmaterial has been deposited on the filter, liquid reagents areselectively dispensed through the same pipette tip into the collectionwell to treat the retained cellular material, after which the automatedarm assembly discards the pipette tip used to dispense the sample fluid,and retrieves a new pipette tip to draw and dispense liquefied paraffininto the collection well to embed the retained cellular matter inparaffin. The paraffin is allowed to cool (and preferably affirmativelychilled), and the cassette is then removed from the cell processor. Thefilter assembly is removed from the cassette, leaving theparaffin-embedded cellular material attached to and sticking out thecollection aperture of the main cassette body. In may be preferable tochill the already solidified paraffin in order to thermally contract theparaffin prior to separately the filter assembly. The embedded cellularmaterial (while still attached to the cassette) is then placed atop of(in direct contact with) an additional piece of paraffin in a thermallyconductive mold configured to seat the main cassette body. The mold isthen heated to soften and at least partially blend together theembedding paraffin and additional paraffin, without being heated to thepoint of softening or liquefying the embedding paraffin such that theretained cellular material therein breaks apart and disburses throughthe embedding paraffin. The mold is then quickly and controllably cooledto bond the “additional” and embedding paraffin.

Other and further aspects and embodiments of the disclosed inventionsare described in the detailed description of the accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

It will be appreciated that the system and apparatus shown in thedrawings are not necessarily drawn to scale, with emphasis instead beingplaced on illustrating the various aspects and features of theillustrated embodiments, in which:

FIG. 1 is a perspective view a cell block preparation system including aprocessing station, a cell block cassette with detachable filter, and afinishing station, constructed according to exemplary embodiments of thedisclosed inventions.

FIG. 2 is a front perspective view of the processing station, with itscabinet doors open to reveal its interior compartments and contents.

FIG. 3 is a side-rear perspective view of the processing station.

FIG. 4 is a partial perspective view of process compartment of theprocessing station.

FIG. 5 is a partial perspective view of a waste compartment of theprocessing station.

FIG. 6A is a partial perspective view of a reagent compartment of theprocessing station.

FIG. 6B is a perspective view of a reagent holder tray that is housed inthe reagent compartment.

FIG. 6C is partial perspective view of a reagent container top and lid,illustrating the outflow and inflow tubing connections.

FIG. 7 is an exploded perspective view of the cell block cassette andfilter assembly.

FIG. 8 depicts a user attaching the filter assembly to the cell blockcassette prior to creation of a new cell block.

FIG. 9 depicts a user detaching the filter assembly from the cell blockcassette following creation of a new cell block.

FIGS. 10A-10C show perspective views of the heated paraffin bathassembly components in the cell processing station.

FIGS. 11A-B depict a user loading a cell block cassette and attachedfilter assembly into the cell block interface prior to creation of a newcell block.

FIG. 12 is a perspective view of the finishing station.

FIGS. 13A-13C depict placement of an additional paraffin block and cellblock cassette in a thermally conductive mold for thermal treatmentusing the finishing station.

FIG. 14 shows the user interface on the finishing station.

FIG. 15 is a flow chart of the process the cell block processing stationundertakes to create a new cell block.

FIG. 16 is a flow chart of the system initialization process for thecell block processing station when undertaking to create a new cellblock.

FIGS. 17A-B is a flow chart of the sample fluid delivery process usedfor creating a new cell block.

FIG. 18A is a flow chart of the sample sipping process in the fluiddelivery process of FIGS. 17A-B.

FIG. 18B is a flow chart of the sample fluid level lowering process inthe fluid delivery process of FIGS. 17A-B.

FIGS. 18C-D is a flow chart of the sample fluid level lowering processin the fluid delivery process of FIGS. 17A-B.

FIG. 19 is a flow chart of the reagent delivery process in the fluiddelivery process of FIGS. 17A-B

FIGS. 20A-B is a flow chart of the paraffin delivery process used forcreating a new cell block.

FIG. 21A-E are respective perspective, top and side views of anotherembodiment of a cell block processing station constructed according toembodiments of the disclosed inventions.

FIG. 21F-J are respective perspective, top and side views of yet anotherembodiment of a cell block processing station constructed according toembodiments of the disclosed inventions.

FIG. 22A depicts a base frame of the processing station of FIGS. 21A-E.

FIG. 22B depicts a base frame of the processing station of FIGS. 21F-J.

FIGS. 23A to 23D are perspective representations embodiments of anautomated arm assembly for use in the cell block processing stationembodiments disclosed herein.

FIGS. 24A and 24B depict one embodiment of a pipette tip connector foruse with the automated arm assembly.

FIG. 25 is a perspective representation of one embodiment of a samplevial interface (or holder) block.

FIG. 26 is a perspective representation of one embodiment of a pipettetip removal post.

FIG. 27 is a representation of an embodiment of a pipette tip removalknife.

FIG. 28 is a representation of another embodiment of a pipette tipremoval post to which a pipette tip removal knife is attached.

FIG. 29 is a representation of an embodiment of a heatsink.

FIG. 30A is a representation of an embodiment of a sample platform.

FIG. 30B is an exploded representation of an embodiment of a heat enginewith a sample platform.

FIG. 30C is a representation of another embodiment of a sample platform.

FIG. 31 is a representation of one embodiment of a heatsink top plate.

FIG. 32 is a representation of one embodiment of a cassette holder.

FIG. 33 is a representation of one embodiment of a clamp plate.

FIG. 34 is a representation of one embodiment of a sensor arm.

FIG. 35 is a representation of a waste evacuation system according toone embodiment that includes a heated valve and a heated reservoir.

FIG. 36 is a further representation of a waste evacuation system

FIG. 37 is a more detailed representation of a heated valve.

FIG. 38 is a representation further showing heated reservoir and heatedvalve components of an evacuation system.

FIG. 39 is a representation of a feedback system to control thepositioning and rotation of a valve of a waste evacuation system.

FIG. 40 is a representation of another waste evacuation system thatseparates and separately processes liquid waste and solid wastegenerated during cell block processing.

FIG. 41 is an exemplary schematic block diagram of a vacuum and pressuredelivery system employed in the waste line of the cell block processingstation.

DETAILED DESCRIPTION OF THE ILLUSTRATED EMBODIMENTS

Systems and methods of embodiments of the present invention providesubstantially automated creation of paraffin-embedded cell blocks byemploying an automated arm in conjunction with a controlled vacuum todeposit a layer of cellular material (e.g., cervical cells obtained forma typical Pap smear) on a removable cell block cassette filter,infiltrate the deposited cellular matter with stain, reagents andparaffin, and then encapsulate the embedded cellular matter withadditional paraffin.

FIG. 1. depicts the main components of an exemplary cell blockprocessing system 20, including a cell block processing station 10, atwo-piece cell block cassette 11 (including a main cassette body 13 anda detachable filter assembly 14) which captures the captures thecellular material and guides infusion of the reagents and paraffin, anda finishing station 12 for encapsulating a newly created cell block inadditional paraffin in preparation for later cutting and slidepreparation. As seen in FIG. 2, the cell block processing station 10includes a reagent compartment 16, process compartment 17 and wastecompartment 18 provided in a single housing cabinet 15. A controller(not shown) including a computer processor and associated memory ishoused within one side of the process compartment 17, and is operativelycoupled with a user interface touch screen 19 located on a frontexterior surface of the cabinet 15. the user interface is preferablyprovided in multiple possible languages and formats, as is well-known inthe art of user interfaces.

Referring to FIG. 3, the back side of the processing station cabinet 15is preferably ventilated 20 to release heat generated by the interiorelectronics. A main power on/off switch 21 is located on the back sideof the cabinet, although other locations may be used, if so desired.Respective ethernet and USB ports 22 and 23 are also provided on theback side of the cabinet 15. The interior cabinet chambers arepreferably fume ventilated using a charcoal-activated filter, which maybe accessed through the filter cover piece 24 on the back of the cabinet15.

Referring to FIG. 4, the process compartment 17 generally includes acell block cassette interface 26 configured for removably seating thecell block cassette and (attached) filter assembly 14. A fluid levelsensor 25 is mounted in a fixed position relative to the cassetteinterface 26 to detect a fluid level in the collection well of the cellblock cassette 14 (described in greater detail herein). In particular,where an ultrasound sensor is used, the sensor 25 is mounted directlyabove and orthogonal to the fluid surface in the collection well. Thesensor 25 is preferably mounted on a pivoting attachment that can bemoved to allow for inserting and removing a cell block cassette from thecell block cassette interface 26. A heated paraffin reservoir 27 (e.g.,containing Paraplast-Xtra® paraffin wax) is disposed to one side, and asample vial interface (or holder) 33 is disposed to another side,respectively, of the cell block cassette interface 26. The sample vialinterface 33 defines a well 35 for removably seating a standard fluidsample vial (not shown) from which cellular matter is to be aspiratedfor creating a respective cell block. The sample vial interface 33 isflanked on its sides by a sample pipette tip holder 32 and a liquidwaste port 34, respectively.

An automated arm assembly 37 is mounted along a rear portion of theprocess compartment 17, and includes a delivery arm 28 that may movedhorizontally along rail 36 and vertically along rail 38. The deliveryarm 28 is coupled to a pipette holder adapted to selectively retrieve,carry and discard (using the pipette tip remover 31) pipette tips 30used for fluid and paraffin aspiration and dispensing during cell blockprocessing. A pipette tip sensor 29 is fixed to the delivery arm 28 fordetecting whether a pipette tip is connected to the tip holder. Thedelivery arm 28 and pipette holder are preferably configured so thatpipette tips attached to the holder will approach the sample collectionwell of a mounted cell block cassette at an angle from the perpendicular(e.g., 15-20°) in order to avoid interfering with the fluid level sensor25 when dispensing sample fluid, stain, reagents and/or paraffin intothe cassette collection well.

Referring to FIG. 5, the waste compartment 18 generally includes a fluidwaste chamber 39 underlying the cell block cassette interface (FIG. 4),and a pipette tip waste bin 40 underlying the pipette tip remover 31(FIG. 4). As will be explained in greater detail herein, the wastechamber 39 is sized for single cell block processes and maintains aclosed system in conjunction with a cell block cassette seated in thecassette interface. As explained in greater detail herein, the contentsof the waste chamber 39 are evacuated into a large waste container (notshown) prior to (or following) each new cell block process.

FIGS. 10A-10C show perspective views of the components of the heatedparaffin bath 27, which includes a main body 64 defining a heated waxtank 68 that is covered using a removable cover plate 66 (FIG. 10A;shown assembled on FIG. 10C). The cover plate 66 reduces the likelihoodthat the wax may become contaminated by airborne pollutants. Prior toprocessing a cell block, pieces of wax paraffin are placed in the heatedwax tank 68 and melted. A temperature sensor 70 is provided for thecontroller to monitor and regulate the temperature of the wax bath 68.The wax bath is divided by a screen 72, with a portion 74 of the bath 68left exposed by the cover plate 66, allowing for a pipette tip carriedby the pipette holder on the automatic arm assembly to be positioned inthe bath portion 74 for aspiration of liquefied paraffin there from. Newpieces of wax are preferably inserted in the portion of the bath 68 thatis underlying the cover plate 66, and the screen 72 keeps any solidpieces that have not yet melted from clogging up a pipette tip submergedin the exposed portion of the bath 74. As described herein, the paraffinaspiration and dispensing process will typically require two separatepipette tips. For this purpose, a pair of pipette tip holding slots 76(FIG. 10B) are built into the paraffin bath structure 27 for holding arespective pair of pipette tips 82 (FIG. 10C).

Referring to FIG. 6A, the reagent compartment 16 stores three separatefluid containers. The first container 41 stores alcohol for use as adrying agent (e.g., 98% isopropyl alcohol, HPLC-UV grade). The secondcontainer 42 stores xylene for use as a clearing agent to remove thealcohol (e.g., reagent grade ACS, histology lab grade 100% xylene). Thethird container 43 stores a staining agent (e.g., eosin Y intensifiedstain available from Fisher Scientific), which is used (optionally) tostain the cellular matter prior to application of the alcohol and xylne.While technically not itself a “reagent”, the stain container 43 is keptin the reagent compartment 16. As shown in FIG. 6B, the fluid containers41, 42 and 43 are seated in respective wells formed in a holding tray 44for added stability. To reduce the chances of confusion, the connectorfeed lines 47 for the fluid containers are preferably different colorsfrom one another, for example, the eosin stain has a red feed line,xylene a green feed line, and the isopropyl alcohol a blue feed line.

As shown in FIG. 6A, possible confusion or entanglement of the fluidfeed lines 47 is further reduced by providing a manifold 48 to keep thelines spaced apart within the reagent compartment 16. The fluidcontainers 41, 42, 43 are provided with respective air inlets (or airinlet lines if they are pressurized) to replace the fluid contents asthey are dispensed. By way of example, FIG. 6C shows a container lid forthe xylene container 42, which includes a green outlet (feed) line 45,and a white air inlet line 46. The respective fluid feed lines 47 areeach coupled to the pipette tip holder, with a respective valve operableby the controller to place the respective individual fluid feed linesfeed line in fluid communication with a pipette tip carried by thepipette holder. In the illustrated embodiment, the eosin stain andxylene feed lines are pressurized using standard in-line pumps (notshown), and the isopropyl alcohol feed line is coupled with an in-linesyringe pump (not shown), which allows the alcohol feed line toalternatively be used as an sample fluid and paraffin aspiration suctionsource by the pipette holder.

Referring again to FIG. 1, as well as to FIG. 7, the cell block cassette11 includes a main cassette body 13 made of molded plastic and having acollection aperture 50 formed therein, and a filter assembly 14removably attached to the main cassette body 13 and defining a samplecollection well 54 in communication with the collection aperture 50 whenthe pieces are attached. A filter 60 (e.g., a “track edge” filtermembrane) is positioned across a bottom surface of the collection well54, and is sized configured to retain cellular matter (e.g., cervicalcells) carried in a sample fluid (e.g., preservative solution) that isdispensed into the collection well 54 and flows across the filter 50. Ithas been found by the inventors that a filter having a relatively highdensity (number per area) of pores having diameters less than about 5microns and, in particular, approximately 3 microns each are suitablefor cell block processing using embodiments of the invention, since muchof the cellular matter and other objects in the sample solution isbetween 7-9 microns in diameter, and larger pores tend to become cloggedprior to when an adequate cell layer has been deposited on the filter.

Still referring to FIGS. 1 and 7, the filter assembly 14 comprises asingle-piece housing or body 80 having a base portion 52 forming thesealing surface, and a neck portion 53 extending from the base portion52. The neck portion 53 defines a perimeter 55 of the collection well54, and has a top end that makes an interference fit with an annulargrove 84 located in a bottom surface of the main cassette body 13 inorder to detachably attach the filter assembly 14 to the main cassettebody 13. The filter assembly housing further comprises a filter supportretention portion 56 extending from an underside of the base portion 52,with a thermally conductive porous filter support member, e.g., asintered metal disc or other thermally conductive porous support 59(such as a metal screen or solid thermally conductive disc with holespunched in it) retained therein, the filter 60 being positioned atop thefilter support member 59 within the collection well 54. The support 59also serves to transmit heat and cooling to the contents of thecollection well via electronics (e.g., a peltier system) located in oraccessed through the waste chamber (not shown). The base portion sealingsurface slopes at a downward angle from the neck portion, such that anoutermost edge 57 of the sealing surface extends to (or beyond) anoutermost edge 58 of the filter support retention portion 56 in order toform a seal with the waste chamber interface (described below) on theprocessing station 10.

In particular, cells must be contained on the filter 60 and the samplefluid, stain, reagents and paraffin must be allowed to pass through thesystem and be discarded in the waste chamber, leaving only cells andhardened paraffin wax behind. As mentioned above, a vacuum in fluidcommunication with the waste chamber interior is applied by thecontroller across the filter 60 at varying strengths (described herein)in order to achieve this fluid pass. Pressured air may also appliedunder certain conditions from the underside of the collection well toensure the pores of the filter membrane are not blocked. The filtermembrane 60 must be able to withstand all of these conditions; and thehousing 80 provides a container for this purpose. The filter may be heatstaked across a bottom area of the (polyester) housing 80 to define turnthe collection well 54. The filter support member 59 (in one embodiment,a sintered bronze or other metal disc) provides a porous, thermallyconductive support to be pressed against the filter membrane 60 tosupport both during vacuum and pressured air cycles.

The housing (or filter holder) 80 and attached (heat staked) filter 60define the collection well 54 for the various liquids and wax to form ahardened protrusion from the mating cassette body 13. This filter allowsthe fluids to pass through but leave cells to collect on the surface ofthe membrane. The vacuum is involved in the process to pull fluidthrough the filter, so a metal or other hard support member 59 is usedto keep the filter 60 from being pulled into the waste chamber by thevacuum. This support member 59 has a given porosity, such as 80 microns,to allow fluid to easily continue to pass through the filter assemblyafter passing through the much smaller filter pores. The sealing surfaceof the base portion 52 forms a vacuum seal on the outer sealing edge ofthe waster chamber interface when the cassette is seated (and latchedinto) the cassette interface in the processing station. The compliantnature of the outer plastic “mushroom” shape of the sealing surface 52provides both adequate vacuum sealing and a proper preload whencompressed to withstand the pressure applied to the under surface duringback pulsing (pressurized air pulsing). The sealing surface of the base52 also provides the system operator with a graspable means to removethe filter assembly 14 from the main cassette body 13, thus leavingbehind a solid protrusion of paraffin wax containing a layer of cells atthe very top end of the wax protrusion when the cell block has beenprocessed. For purposes of illustration, FIG. 8 depicts an operatorattaching the filter assembly 14 to the main cassette body 13 prior toprocessing a new cell block. FIG. 9 depicts the operator detaching thefilter assembly 14 from the cell block cassette 13 following creation ofa new cell block 62.

Preferably, the main cassette body has a substantially planar topsurface and a substantially planer bottom surface, and the top andbottom surfaces are substantially parallel with one another, to allowfor ease in stacking of multiple cassette bodies having their respectivefilter assemblies removed (e.g., for labeling, bar code application,etc).

In alternative embodiments, many other structures and methods ofobtaining the same outcome of creating a collection well are possible.For example, a radial seal can be used to create the vacuum seal,something similar to the seals used in syringes. This would require theuse of a more compliant material, and possibly with a form of rubberinsert as a seal. A face seal also could be used for forming the vacuumseal, which would include the use of some sort of gasket or similarmaterial either free to float on the instrument or fixed to theinterface. The gasket material could also be placed on the base of thefilter holder depending on the design. Other shapes and geometries canbe used for the filter housing 80, which would create various shapes andsizes of the collection well. One such alternative shape is oval, whichwould reduce the need for a re-melt step (discussed in conjunction withthe finishing station 20), since the microtome would be able to make aribbon out of the block as is.

For purposes of illustration, FIGS. 11A and 11B depict a user loading acell block cassette and attached filter assembly 11 into the cell blockinterface on the processing station prior to creation of a new cellblock. Notably, a clamping member 79 of the sensor assembly 81 may bepivoted and moved aside (FIG. 11A) for inserting the cassette 11 intothe interface, after which the clamping member is secured to theinterface (FIG. 11B) so that the sensor assembly 81 (and sensor 25) areproperly aligned with the collection well of the (now secured) cellblock cassette. Clamping down on the cassette and filter assembly 11also secures the vacuum seal formed between the sealing surface 52 ofthe filter assembly and the waste chamber interface (not shown). This isneeded in particular to avoid displacement of the cassette/filterassembly 11, or failure of the seal due to backpressure maintained inthe vacuum chamber 39 to prevent fluids from passing through the filter60 unless the vacuum is ON, as well as the more significant backpressure that may be used for temporarily lifting the collected matteroff the filter (discussed in detail below). At the time the cell blockcassette is loaded into the cassette interface, the user should alsoverify that the fluid level sensor is clean and unobstructed.

When dispensing the sample fluid into the collection well, certainsamples will tend to clog the filter before a sufficient cellularmaterial layer is retained. Such samples usually contain smallindividual cells, such as lymphocytes and other inflammatory cells,which have a tendency to stack and almost immediately impede the fluidflow across the filter. In order to collect additional cellularmaterials, and therefore a larger cell layer retained by the filter, airbubbles are pushed (or pulsed) through the filter from the waste chamberto temporarily lift the cellular materials away from the filter surface,and allow more liquid to pass through. This is done by applying apressure within the waste chamber just larger than the bubble point ofthe filter material to gently lift the cellular material from the filtersurface. FIG. 41 shows embodiment for the vacuum and pressure deliverysystem for the waste chamber. The pressure pulse valve allows a smallpressure to be injected in the waste chamber. This small pressure isjust larger than the bubble point of the membrane filter. When samplecollection has stalled, this small pressure pulse can lift the samplefrom the filter and allow more sample to be collected.

In alternate embodiments, another form of sample fluid agitation thatmay enhance the amount of sample collected is simply mechanicalvibration of the cell block cassette and filter assembly 14. Suchmechanical motion would likely prolong the amount of time it takes tocompletely clog the filter pores, thereby allowing more sample to becollected. One or both of lateral vibration (parallel to the filtermembrane) and vertical motion may be used. The frequency of verticalvibration would appear to be more sensitive to the amount of sample(mass) present in the collection well. Actuation of the mechanicalvibration of the cell block cassette can be accomplished in a verity ofways. For example, a custom piezoelectric actuator designed into theengine assembly could be used to deliver the proper energy required tomaximize cellular matter collection. A voice coil design may also beable to deliver enough low frequency energy to enhance sample fluiddelivery. Another option would be to deliver energy into the cel blockcassette through an acoustic pressure wave induced into the wastechamber interior below the filter membrane. The acoustic energy wouldvibrate the filter membrane during sample collection, thus, keeping thepores open longer.

As is described further in conjunction with the cell block process flowcharts of FIGS. 15-20, when dispensing the sample fluid into thecollection well, certain samples will tend to clog the filter before asufficient cellular material layer is retained. Such samples usuallycontain small individual cells, such as lymphocytes and otherinflammatory cells, which have a tendency to stack and almostimmediately impede the fluid flow across the filter. In order to collectadditional cellular materials, and therefore a larger cell layerretained by the filter, air bubbles are pushed (or pulsed) through thefilter from the waste line to temporarily lift the cellular materialsaway from the filter surface, and allow more liquid to pass through.This is done by applying a pressure within the waste line just largerthan the bubble point of the filter material to gently lift the cellularmaterial from the filter surface.

FIG. 41 shows embodiment for the vacuum and pressure delivery system forthe waste chamber. The pressure pulse valve allows a small pressure tobe injected in the waste chamber. This small pressure is just largerthan the bubble point of the membrane filter. When sample collection hasstalled, this small pressure pulse can lift the sample from the filterand allow more sample to be collected.

An alternative form of sample agitation that appears to enhance theamount of sample collected is simply mechanical vibration of thecassette/filter. The mechanical motion prolongs the amount of time ittakes to completely clog the filter pores allowing more sample to becollected. Lateral vibration parallel to the filter membrane appears towork better than vertical or perpendicular motion. The frequency ofvertical vibration also appears to be more sensitive to the amount ofsample (mass) present. Actuation of the cassette/filter can beaccomplished in a verity of ways. A custom piezoelectric actuatordesigned into the engine assembly could deliver the proper energyrequired to maximize sample collection. A simpler voice coil design mayalso be able to deliver enough low frequency energy to enhance sampledelivery. Another option would be to deliver energy into thecassette/filter through an acoustic pressure wave induced into the wastechamber volume below the filter membrane. The acoustic energy wouldvibrate the filter membrane during sample collection keeping the poresopen longer.

FIGS. 15-20 are detailed flowcharts illustrating the processesundertaken by the processing station 10 when processing a cell block.Notably, the processor of the processing station keeps a log of the cellblock events, which may be printed or transferred using the Ethernet orUSB connections.

In particular, FIG. 15 is a flow chart of a cell block process 800 usingthe system 20 of FIGS. 1-14. The user has already loaded a new (andsterile) cell block cassette and filter assembly into the cassetteinterface of the processing station (FIGS. 11A and 11B), with thesealing surface 52 of the filter assembly forming a vacuum seal with theinterior of the waste chamber 39. The user has also verified that thereis adequate paraffin in the heated wax bath 68 (FIGS. 10A-C), andsterile pipette tips available at the respective sample interface andwax bath. The amount of sample fluid that is used for processing a cellblock is normally limited to 20 ml, but may less may be used if asufficient cell layer is collected by the filter with less sample fluid.Thus, a user should verify that the sample vial has at least 20 ml offluid prior to commencing the cell block process 800.

The cell bock process 800 starts with an system initialization cycle 802of the processing station, which (referring also to FIG. 16) includesevacuating the waste chamber (830) any liquid or solid waste remainingfrom the last cell block process (discussed below) by opening a heatedwaste chamber evacuation valve; testing the heating/cooling system (832)for the sample collection well (i.e., the peltier system thatcontrollably heats and cools, respectively, the filter support 59, whichin turn heats or cools the contents of the collection well 54); andpriming the isopropyl alcohol feed line (834) using the liquid wasteport provided at the sample vial interface to dispose of the alcohol.The system then loads a sample fluid pipette tip (836) and tests thefluid level sensor and waste chamber vacuum systems (838). Assuming noerrors are encountered during the initialization process 802, theinitialization is completed and the sample fluid aspiration processcommences. If one or more errors occur during the initialization, thesystem reports these to the user (835) and the cell block process iscancelled (833).

Referring back to FIG. 15, the user indicates whether the cell blockprocess will be fully or “partially automated” (804), where “partiallyautomated” indicates that the user will be manually loaded tissuefragments into the collection well in addition to the sample fluidaspiration. If “partially automated” is selected, the system pauses(806) to allow the user to manually insert (e.g., with tweezers or thelike) tissue fragments into the collection well. For example, this maybe required because there are tissue fragments in a patient sample thatmay otherwise clog the aspiration pipette tip. In any event, once thetissue fragments are manually placed in the collection well, the systemcontinues the process (809). Notably, there may be differences in theautomated processing if the user has manually inserted tissue fragments,e.g., pressurized air back pulsing (or “burping”) through the filter isnot used, and the amounts and exposure times of the respective reagents(isopropyl alcohol and xylene) and paraffin are much greater than if thecellular materials are limited to those collected through sample fluidaspiration. Also, in order for the system to continue with the automatedcell block processing, a fluid aspiration component must still be used(and a sample fluid vial is required), even if the tissue sample islimited to a relatively large fragment taken from a core biopsy.

The system then undertakes the process (810) of aspirating sample fluidfrom the sample vial and dispensing the sample fluid into the collectionwell of the cassette-filter assembly using a pipette tip. The amount offluid that is dispensed at any one time always depends on the existingfluid level in the collection well (measured by the sensor 25) and mayalso depend on the most recently measure flow rate across the filter.Also, the waste chamber back pressure is maintained so that no fluidflows across the filter unless the vacuum is activated. Because thepipette tips have a fixed volume, and the syringe pump on the isopropylalcohol feed line may be precisely controlled, the controller cancarefully track how much fluid is aspirated, from the sample vial, howmuch has been dispensed into the collection well, and how much remainsin the pipette tip at any given time. Prior to dispensing fluid into thecollection well (including the first time), the controller checks on thefluid level in the collection well (840). Depending on the amount offluid already in the collection well, the controller then activates thesyringe pump on the alcohol feed line to dispense a limited volume ofsample fluid into the collection well (842). The details of this sample“sipping” process 842 are shown in FIG. 18A.

In particular, with reference to FIG. 18A, the sample sipping process842 starts with the controller calculating the available fill volume 866in the collection well based on the fluid level sensor reading. Then,depending on the current volume, if any, of sample fluid remaining inthe pipette tip from a previous sipping process, the controllerdetermines whether additional sample fluid needs to be aspirated intothe pipette tip (868) in order to meet the calculated fill volume. Ifadditional sample fluid is needed, the controller first dispenses theremaining sample fluid in the pipette tip into the collection well(870), and then aspirates additional sample fluid from the sample vial(872) and dispenses the balance of the fill volume into the collectionwell (874) to complete the sip process (875).

After the sample fluid is dispensed into the collection well, thecontroller activates the vacuum pump (844) in fluid communication withthe waste chamber sample to draw the fluid across the filter, whilemonitoring the fluid level (846) in the collection well as a function oftime. The details of the “lower sample level” process (848) are shown inFIG. 18B. In particular, two threshold items are monitored: a pulsetarget flow rate, which is the flow rate at which a back pressure (or“burping” pulse) may be applied, and a low threshold fluid level, whichis the threshold fluid level that is maintained in the collection wellat the end of a sample sip process. To initiate the “lower sample level”process (848), a “timeout” timer is started by the controller (870), andthe fluid level is monitored with respect to the timer (871). From thisinformation, along with the known dimensions of the collection well, aflow rate across the filter may be calculated (872). So long as thesample fluid drains down to the threshold level (874) without any“timeout”, then the process is complete, and the flow rate is noted bythe system. If the sample level does not fall below the threshold, butthe flow rate is less then the target rate, then the process iscomplete. However, the sample level does not fall below the thresholdand the flow rate is not less then the target rate, then the monitoringof the fluid level and flow rate continues (steps 871-872 repeated). Inone embodiment, the fluid level and flow rate are monitored every 20milliseconds for up to 90 seconds, after which, the system “times out”and an error message is provided to the user (876).

Once the fluid level in the collection well is lowered to a minimumlevel (preferably at least some fluid is left in the collection well sothat the retained cellular matter is not exposed to the air), the vacuumis turned OFF (850). The flow rate of the just-completed sample sip isthen compared with a target flow rate (851). If the flow rate wasgreater than the target flow rate, and if the system did not “time out”while monitoring the fluid level (856), and if less than 20 ml of samplefluid have been dispensed (858), a new sample sip is commenced byreturning to step 840. However, if the flow rate on the just finishedsip was less than the target rate (851), or the system “timed out” whilemonitoring the fluid level (852), and if the cell processing was not“partially automated,” (853, then the system undergoes a lower fluidlevel process (856) using back pulse of air pressure supplied from thewaste chamber in order to momentarily lift the cellular matter off thefilter and allow more sample fluid in the collection well to passthrough. The details of the “lower fluid level” process (854) are shownin FIGS. 18C-D.

In particular, the lower fluid level process 854 begins by reading thefluid level in the sample collection (880) to ensure the level is highenough to turn ON the vacuum in the waste chamber to start drawing fluidout of the collection well (881). If the fluid level is not high enoughto turn ON the vacuum, the lower fluid level process 854 is considereddone. If there is sufficient fluid level to draw some down through thefilter, then the vacuum is turned ON (882), and the system notes thetime and fluid level (884) and monitors same every 20 milliseconds untilthe process is completed or times out. If a back pulse is “requested”(885), meaning the cell block process is fully automated and the fluidis sample fluid, the system next checks to make sure the fluid level isnot too high for back pulsing (886), since otherwise the pressurized aircoming up through the filter from the waste chamber can cause the samplefluid to splash out of the collection well. The controller also verifiesthat a previous back pulse was not administered within a predeterminedinterval (887) to ensure that back to back pulses are not administered.If the fluid level is not too high, and a previous back pulse has notbeen administered within the allowed interval, then the vacuum is turnedOFF (888), and the source of pressurized air is put in communicationwith the waste chamber at a pressure sufficiently high to create apressurized air back pulse (or “burp”) (889) through the filter to liftthe cellular matter away from the filter surface. The pressurized air isthen turned OFF, and the vacuum turned back ON (890), to allow for thefluid to pass across the momentarily cleared filter.

Following application of a back pulse, the fluid level and flow ratecontinues to be monitored in the same manner as if no back pulse hadbeen applied (in 20 ms cycles). The controller evaluates whether a“target level” is reached (892), which is based on a composite test thatfactors in both whether the fluid level is within a threshold of thelower limit (e.g., within about 10% of the total collection well heightor volume) and the process has taken longer than a specified amount oftime (e.g., approximately 40 seconds), in which case it is assumed thatthe system has pulled as much sample fluid through the filter as isneeded, and the vacuum is turned OFF (895) the lowering fluid levelprocess 854 is complete. On the other hand, if this target level is notreached after 90 seconds of processing time (894), and the process isended with the user received a system error message (896), and the cellblock processing ceases for that sample.

Returning to FIGS. 17A-B, if back pulsing through the filter allows thefluid to drain from the collection well prior to a “time out” (856), andif less than 20 ml of sample fluid have been dispensed (858) and theflow rate is lower than the target flow rate (860), then the vacuum ratemay be increased (864) (e.g., from 10% to 50% following an initialsample sip) and a new sample sip is commenced by returning to step 840.Notably, in embodiments of the invention, the vacuum rate is not raisedhigher than 50% of maximum for pulling sample fluid. However, if despiteback pulsing through the filter the fluid does not drain from thecollection well prior to a “time out” (856), or if 20 ml of sample fluidhave been dispensed (858), or if the flow rate drops below the targetflow rate (860) at the full vacuum strength applied for pulling samplefluid (862), then no more sample fluid is dispensed into the collectionwell.

Returning to FIG. 15, once the aspiration and dispensing of the samplefluid is complete, eosin stain is (optionally) injected into collectionwell (812) by the automated arm assembly. In particular, the controllercloses the valve on the alcohol feed line at the pipette tip holder andthe opens the valve on the eosin feed line. Because the eosin containeris pressurized, the eosin self-dispenses through the same pipette usedfor the sample fluid into the collection well, and the automated armneed not be repositioned. Once the desired amount of eosin stain hasbeen dispensed, the controller closes the valve on the eosin stain feedline, and selectively re-opens the valve on alcohol feed line, and thesyringe pump is activated to meter out the isopropyl alcohol (a dryingreagent to displace the water content from the cellular mater) into thecollection well according to the reagent delivery process (814)illustrated in FIG. 19.

In particular, the controller selects the reagent (in this case, thealcohol), starts a timer (900), and reads (detects) the fluid level ofthe remaining sample fluid in the collection chamber (902) in order tocalculate how much alcohol to initially dispense (903) withoutoverflowing the collection well. The calculated amount of reagent(alcohol) is then dispensed (via precise operation of the syringe pump)into the collection well (904) and the fluid level is again read (905)to verify the actual amount that was dispensed (906). The timer is thenread (907), followed by a calculated ouase time (908) and delay pausetime (910). Essentially: the controller calculates the amount of time towait before passing the reagent through the filter in order toadequately expose the cellular material to the respective reagent. Ifthe flow is fast, the total time needed to fill and empty the collectionwell such that the volume is dispensed may be less than the specifiedexposure time. So, the system preferably delays a bit in each reagentdispense process to compensate. Roughly, the exposure times the numberof fills minus the time to lower level in the last iteration.

The fluid level is read again (912) and the lower fluid level process854 (FIGS. 17A-B) is performed, except that no back pulsing is done. Atthat point, the controller determines whether the target volume of thereagent (alcohol) has been dispensed, and for the required exposure time(916). If so, then the reagent delivery process is done. If not, theprocess returns to step 902 and is repeated.

Once the alcohol has been fully dispensed, the controller closes thevalve on the alcohol feed line, and selectively opens the valve on the(pressurized) xylene feed line, as the reagent delivery process (814) isrepeated for dispensing xylene (a clearing reagent to eliminate thealcohol). For a fully automated process, preferably 3.0 ml each ofisopropyl alcohol and xylene are respectively dispensed, with at least90 seconds of exposure of the cellular material to each regeant in thecollection well. For a semi-automated process (meaning some cellfragments are manually introduced into the cell collection well), 15 mlof alcohol for at least 5 minutes exposure is followed by 20 ml ofxylene for at least another 5 minutes exposure. In either case, theremaining xylene is vacuum pulsed out of the collection chamber justprior to when the paraffin is first dispensed therein.

Returning to FIG. 15, with a small portion of the xylene still in thecollection well to prevent the cell layer from direct air exposure, thecollection well is heated (818) via heating the metallic support member59 underlying the filter 60. The reason for heating the collection wellis to prevent the liquefied paraffin from immediately solidifying uponcontact. The automated arm disposes of the pipette used for the samplefluid, stain and reagents, and connects to a new pipette tip at the waxstation. Using a separate pipette tip for aspirating the paraffin thanwas used for aspirating the sample fluid helps avoid cross-contaminationbetween cell samples. For this same reason, a separate pipette tip ispreferably used each time additional paraffin is aspirated from the waxbath.

The process for delivery of the paraffin (820) is illustrated in FIGS.20A-B. In particular, a new pipette tip (located at the wax bathstation) is secured to the pipette tip holder (920), and the controlleragain reads the fluid level in the collection well (921). As the newlysecured pipette is dipped into the wax bath and loaded with a firstvolume of the liquefied paraffin, the waste chamber vacuum is switchedON (full strength) to remove the residual xylene through the filter andthen switched back OFF (922). The liquefied paraffin is then dispensedinto the collection well (924), and the vacuum is switched back ON (925)to draw the liquefied paraffin into the cellular matter, through thefilter pores and into the waste chamber, to thereby fully embed thecellular matter that was retained by the filter. The flow of theparaffin is monitored in the same manner as the prior fluids, bytracking the fluid level in the collection well and the elapsed time(927). The level of the paraffin is compared to a desired low level(928) indicating a full absorption of the wax into the cellular matter.Once the desired absorption level is detected by the sensor, the vacuumis switched off (934). In the mean time, if the absorption level is notyet reached prior to the system timing out (930). In the case of a “timeout” on the paraffin level lowering watch, the vacuum is shut OFF.Similarly, a further “time out” (932) is monitored by the controller totrack the time that the liquefied paraffin has been in the pipette tip.Because the paraffin can quickly solidify, preferably no longer than 30seconds is provided for this “paraffin tip” time out period.

Whether because of the level sensed, or a “time out” was reached, oncethe vacuum is shut OFF, the controller again reads the fluid level inthe collection well (935) to determine whether the desired paraffinthrough volume has been reached (936). If so, (or if there was a “timeout” for the lowering paraffin level watch), then the controllerdetermines what liquefied paraffin remains in the pipette tip (944),dispenses same into the collection well (945), and ejects the pipettetip (946). If a further pipette is available (947), the further tip isretrieved by the pipette holder (948), loaded with further liquefiedparaffin from the wax bath (949/950), and dispensed into the collectionwell (952) to fully encapsulate the cellular matter. If at step 936 thedesired paraffin through volume is was not reached, and there was not a“time out” on the lowering paraffin level watch (938) or a paraffin tiptime out (940), then (again), the controller determines what liquefiedparaffin remains in the pipette tip (942), dispenses same into thecollection well (943), and ejects the pipette tip (954). Further steps956 (verifying a second pipette tip is available), 958 (loaded thesecond tip on the pipette tip holder), 959 (calculating what remainingparaffin is needed) and 960 (dispensing same into the collection well),are then performed, as illustrated in FIG. 20B. In the event no secondpipette tip is available, a warning is sounded to the user, and theparaffin delivery process ceased (957).

For a fully automated process, approximately 1.5 ml of paraffin isintroduced and held at temperature to assure the paraffin remains meltedwith an exposure time of 20 seconds. In a semi-automated process, 1.5 mlof paraffin is introduced and kept molten for at least 7.5 minutes.Using separate pipette tips for aspirating the sample fluid and fordispensing reagents versus aspirating the paraffin helps avoidcross-contamination between cell samples. This is the same reason thatseparate pipette tips are used for aspirating paraffin if more than oneaspiration from the wax bath is needed.

Extra care must be taken when dispensing the paraffin to be sure thatthe pipette tips do not become clogged with solidified pieces of wax, orthat solidified pieces are introduced into the collection well, as thismay cause the cell block to later crack or fall apart. Thus, the systemimposes a strict time limit on how long it will allow continueddispensing of paraffin from the same pipette tip. After the paraffin isintroduced and drawn into the cellular material (and through the filter60 and substrate 59) by the vacuum, a further quantity is poured intothe well with the vacuum off, in order to fully embed the cellularmatter. The collection well is then chilled (822 in FIG. 15), e.g., byreversing the peltier system used to heat the substrate 59 so that itnow chills the substrate, to solidify the paraffin in the collectionwell prior to when the operator removes the cassette and newly formedcell block from cassette interface and removes the filer assembly fromthe cassette and paraffin block. In order to separate the filterassembly 14 from the main cassette body 13, it may be advisable to firstchill the already solidified paraffin to cause further thermalcontraction of same. This can be accomplished, for example, by sprayinga cold gas (compressed air) onto the cell block or placing it in afreezer for a short period of time. As the wax contracts, it releasesfrom the respective filter and support member.

FIG. 12 is a perspective view of the finishing station 12, whichgenerally includes a housing 92 for enclosing the processor andelectronics and heat exchanger, a thermally conductive heating/coolingplate 86, a simple user-interface (control panel) 88 (FIG. 14), and aclear plastic cover 90 for the heating/cooling plate 86. As illustratedin FIGS. 13A-13C, the finishing station is used to embed the cell blockin additional paraffin, in particular the end of the cell blockcontaining the cell layer.

In particular, a piece of paraffin 91 is first transferred from itspackaging 89 into the re-melt well 93 of thermally conductive (e.g.,stamped metal) embedding mold 87 (FIG. 13A). The embedding mold 87 isthen on the heating/cooling plate 86 on the finishing station 12 (FIG.13B), and the operator starts the unit using the user interface 88 beginmelting the wax 91. When the embedding wax 91 is completely melted(about seven minutes), the cell block cassette 11 is placed into thewell 93 of the mold 87 by fitting one end into the mold 87 and loweringthe cassette until it is fully inserted into the mold, with the cellblock paraffin side face side and into the liquid paraffin. Preferably,no air bubbles are trapped between the melted paraffin and the cellblock paraffin. The unit then continues to apply heat to the plate 86,until the embedding paraffin on the cell block has softened and startedto melt. At this point, the plate is abruptly switched over to cooling.In particular, it is important that the re-melt process employ be fastand controlled heating, followed by fast and controlled cooling. In oneembodiment, a method for the paraffin re-melt includes placing theparaffin-embedded cellular material atop an additional amount ofparaffin; controllably heating to thereby soften and at least partiallyblend together the embedding paraffin and additional paraffin, withoutsoftening or liquefying the embedding paraffin to a point that theretained cellular material therein breaks apart and disburses throughthe embedding paraffin; and controllably cooling to thereby bond theadditional paraffin to the embedding paraffin.

FIGS. 21A-J depict further embodiments of a cell block processingstation, which are similar if not identical in most aspects to theprocessing station 10 of the above-described system 20, and are beingshown and described herein to provide additional details regardingmaking and using embodiments of the disclosed inventions. In particular,FIGS. 21A-E depict the structures and features of one processing station100, which are assembled on a respective base 101 and base frame 102,shown in FIG. 22A. FIGS. 21F-J depict the structures and features ofanother processing station 150, which are assembled on a base frame 152,shown in FIG. 22B.

A sample arm 104 is connected to the base frame 102 of the processingstation 100 by a robotic arm 106. FIG. 23A shows a perspective drawingof the sample arm 104. The robotic arm 106 is configured to movelaterally along the base frame 102 on a horizontal railing 108. Therobotic arm 106 can also move vertically with respect to the base frame102 along a vertical railing 110. The vertical railing 110 is connectedto the horizontal railing 108. Thus, using a combination of thehorizontal railing 108 and vertical railing 110, the sample arm 104 canmove to various locations within the processing station 100. Inparticular, the vertical railing 110 is preferably connected to thehorizontal railing 108 at a slight angle from perpendicular, e.g.,approximately 15° (so that the vertical railing 110 forms an angle ofapproximately 75% with the horizontal railing 108), in order to allow atip of an attached pipette to pass by a level sensor 124 aligned overthe cassette well. To the sample arm 104 is attached a pipette connector402, as shown in FIGS. 24A-B. The lower end 404 of the pipette connector402 is configured to attach to a pipette, such as a disposable plasticpipette.

A plurality of tubes, not shown, are connected to the upper end 406 ofthe pipette connector 402 when the pipette connector 402 is placed onthe sample arm 104. One of the tubes is a vacuum tube, which is filledwith alcohol (as a source of reagent, the use of which is describedbelow) and connected to a vacuum source, not shown. When a pipette isattached to the pipette connector 402 and the distal tip of the pipetteis immersed in a fluid, the vacuum causes the fluid to flow into thepipette.

Other tubes connected to the upper end 406 of the pipette connector 402cause the pipette connector 402 to be in fluid communication with aplurality of liquid sources 112. The liquid sources 112 hold reagents,such as xylene and alcohol, used for the preparation of cell blocks.

A vial holder block 114 is located in the processing station 100. Thevial holder block 114, also shown in FIG. 25, is configured to hold avial containing a biological sample. The biological sample comprisescells that are to be embedded in wax.

The processing station 100 also comprises a pipette removal element. Thepipette removal element comprises a pipette removal post 116, also shownin FIG. 26, and a pipette removal knife 118, also shown in FIG. 27. Thepipette removal knife 118, which is mounted on top of the pipetteremoval post 116, comprises a flared opening 702. When the pipetteconnector 402, having a pipette attached thereto, is laterally insertedinto the opening and then moved upward, the pipette is then detachedfrom the pipette connector 402. A liquid waste line is provided intowhich the contents of a pipette tip can be disposed.

The processing station 100 includes a sample platform 123. In someembodiments, the sample platform 123 comprises a heatsink 1102 (FIG.29). A top plate 1202 (FIG. 31) covers the heatsink 1102. A cell blockcassette and filter assembly holder 1302 (FIG. 32), is configured to beplaced in the opening 1204 of the top plate 1202. The cassette holder1302 is configured to hold a cassette assembly (e.g., the cell blockcassette 11). A clamp plate 1402 (FIG. 33) is configured to be placedover the cassette holder 1302, such that the opening 1404 of the clampplate 1402 is placed over the opening 1304 of the cassette holder 1302.

A sensor 124 is provided, which measures the level of liquid in thecassette well 1612, for example, using ultrasound wave energy, as isdone in the ML series of ultrasonic sensors produced by Cosense. Thesensor 124 is held in place by a sensor arm 1502 (assembly shown in FIG.34).

The processing station 100 also includes a vacuum source 126. The vacuumsource 126 is connected to the cell block sealing surface 52 on thefilter assembly 14, through an air-tight channel 128 so that it canapply vacuum to the collection well and draw liquid through the filter60. The liquid drawn from the collection well is held in the wastechamber 39. In some embodiments, the vacuum source 126 comprises apressure gauge that can measure the pressure differential between theair over the collection well 1612 and the air-tight channel 128.

FIGS. 21F-I depict the components of processing station 150, which areall attached to the base frame 152. The processing station 150 includesthe same robotic arm 106, horizontal railing 108, and vertical railing110 as station 100, which operate in a similar manner as describedabove.

FIG. 23B shows a sample arm 154 is connected to the horizontal railing108 and the vertical railing 110 of the processing station 150 by arobotic arm 106. FIG. 23C shows a perspective drawing of the sample arm154.

To the sample arm 154 is attached a pipette connector 402, as shown inFIGS. 23C and 23D, through a connector head 351. The lower end 404 ofthe pipette connector 402 is configured to attach to a pipette 352, suchas a disposable plastic pipette, as shown in FIG. 23B. A spring 354dampens the pressure on the pipette connector 402 when a pipette 352 isbeing attached thereto.

To the sample arm 154 are also attached a plurality of valves 356, suchas solenoid valves. A plurality of tubes 358 connect at one end to thevalves 356 and at the other end to sources of reagents 156. A pluralityof tubes, not shown, connect the valves 356 to the upper end 406 of thepipette connector 402 when the pipette connector 402 is placed on thesample arm 154. The tubes, which are a vacuum tube and other reagenttubes, operate in a similar manner as described above.

A vial holder block 164 is located on the processing station 150. Thevial holder block 164 (FIG. 25), is configured to hold a vial in theopening 552. The vial, not shown, typically contains a biologicalsample. The biological sample comprises cells that are to be embedded inwax. The vial holder block 164 comprises holes or openings 554, whichare configured to allow a screw to pass through to fasten the vialholder block 164 to the frame 152. The block 164 also comprises anopening 556 configured to hold a pipette tip. The block 164 furthercomprises a purge opening 558, which in some embodiments is connected tothe waste disposal unit, discussed below. In some embodiments, thesample arm 154 obtains a pipette tip. The distal end of the pipette tipis lowered into the biological sample solution within a vial placed inthe opening 552. Vacuum is applied to the interior of the pipette tip,which causes aspiration of some of the sample solution to enter thepipette tip. In some embodiments, the sample arm 154 removes the pipettetip from the biological solution and transfers it to the purge opening558, where the biological sample within the pipette tip is forced out ofthe pipette tip and into the purge opening 558. The process of obtaininga sample and purging it may be repeated more times. This process lowersthe risk that the biological sample within the pipette tip would becontaminated by any contaminants inadvertently present in the pipettetip.

It is also envisioned that any time the pipette tip is to be rinsed, orthe contents thereof purged, the contents of the pipette tip can beemptied into the purge opening 558. This includes any time the pipettetip is to be washed between the use of the various reagents. Theprocessing station 150 also comprises a pipette tip removal element. Thepipette tip removal element comprises a pipette removal element 166,also shown in FIG. 28. The pipette tip removal element 166 comprises apipette removal post 750 and a pipette removal knife 158. In someembodiments, the pipette removal post 750 and the pipette tip removalknife 158 are molded together as one piece. In other embodiments, thepipette tip removal knife 158 is a separate unit from the pipette tipremoval post 750. In these embodiments, the pipette tip removal knife158 is mounted on top of the pipette tip removal post 752. The pipettetip removal knife 158 comprises a flared opening 752, which operates ina similar manner as the flared opening 702, discussed above. A waxstation (not shown) identical to that shown in FIGS. 10A-C is alsoprovided.

The processing station 100 also comprises a sample platform 173 (FIG.30A). In some embodiments, the sample platform 153 is mounted over athermal engine 1151 (FIG. 30B). A top plate 1152 covers the thermalengine 1151. A waste tube 1170 is placed in an opening 1172 of thethermal engine 1151, which allows for the waste from the preparing thesample in the cassette to flow to the waste disposal unit, discussedbelow. A cassette holder 1164 is configured to be placed in the opening1156 of the top plate 1152, over the waste tube 1170. The cassetteholder 1164 is configured to hold the cell block cassette and filterassembly 11. The cassette holder 1164 is held in place further by ahinged cover 1154. The hinged cover 1154 is attached at one end to ahinge 1158, which in turn is connected to the top plate 1152. At theother end, the hinged cover 1154 is held in place by a knob 1160attached to a screw (not shown). To place a cassette on the sampleplatform 173, the knob 1160 is turn to loosen the screw. The hingedcover 1154 is turned upward about the hinge 1158.

FIG. 30C shows the configuration of the hinged cover 1154, with thesensor 124, discussed below, attached, when the hinged cover 1154 isturned upward about the hinge 1158. The cassette is then placed in thecassette holder 1164. The hinged cover 1154 is then turned downwardabout the hinge 1158 and is secured in place by turning the knob 1160.The sample platform 173 comprises holes or openings 1162, which areconfigured to allow a screw to pass through to fasten the sampleplatform 173 to the frame 152. A sensor 124 is provided, which measuresthe level of liquid in the cassette collection well, for example, usingultrasound wave energy, as is done in the ML series of sensors producedby Cosense, or by a Baumer Ultrasonic Level Sensor with beam columnator,such as UNAM 12U9914/S14D. The sensor 124 is held in place by a sensorarm 11174.

The processing station 150 also comprises a touch screen display 174.The touch screen display 174 is electronically coupled to the othercomponents in the system and allows the user to select which reagents touse, the amount of reagents to be used, the order by which the reagentsare used, and other functional elements of using the processing station150.

A further aspect is directed to a heated waste evacuation system,apparatus and method for controllably and safely evacuating flammableliquid and solid waste that is generated during cell block processing.According to one embodiment, an apparatus for evacuating cell blockprocessing waste includes a heated valve, such as a heated ball valve,that is positioned below a reservoir that collects waste. The valve canbe heated by one or more heating elements, such as cartridge heaters.According to another embodiment, an apparatus for evacuating cell blockprocessing waste includes a combination of heated valve and a heatedreservoir. The heated valve can be heated by one or more heatingelements, such as cartridge heaters. The reservoir can be heated by aheating element, such as a foil heater. The valve can be controllablyopened and closed to allow liquid and solid waste and mixtures thereofto be collected, heated and released. Heating waste components ensuresthat solids, such as paraffin wax, remain molten or partially molten tofacilitate waste disposal in a controlled and efficient manner.

More particularly, referring to FIGS. 35-38, a waste evacuation system2800 includes a reservoir or housing 2802 that defines a wastecollection area 2804 therein. Solid and liquid waste (such as wax,xylene, alcohol and mixtures thereof) pass from a cell block engine2803, through a waste tube, such as waste tube 1170, and into the wastecollection area 2804. Cell block engine 2803 generally refers to theunit or components that generate a cell-bearing block. Waste can becollected from the cell block engine 2803 by gravity and also by vacuum,which can be supplied via vacuum port 2806.

In the illustrated embodiment, the waste tube 1170 includes two tubes—anouter tube 1170 a and an inner tube 1170 b. The bottom edge 2808 of theinner tube 1170 b defines a waste drip-edge 2808. The inner tube 1170 ballows waste from the cell block engine 2803 to drip from the knifeedge, and the outer tube 1170 a prevents falling waste from being drawninto the vacuum. In the illustrated embodiment, the outer tube 1170 aextends downwardly into the reservoir 2802 further than the inner tube1170 b. The tubes 1170 a and 1170 b can be concentrically arranged in adual tube configuration as shown in FIG. 35. The reservoir 2802 thatholds the waste can be, for example, a single body housing made of 6061anodized aluminum.

A heating element 2812 is applied to or around the reservoir 2802 toheat the waste held in the collection area 2804. For example, accordingto one embodiment, a foil heater 2812, such as a Minco foil heater, iswrapped around the reservoir 2802. An undercut can be added to positionthe foil heater 2812 as necessary. The foil 2812 heats solids such asparaffin wax to maintain the solids in a molten or semi-molten state.This prevents wax from building up on the interior walls of the chamberof the valve 2820.

The bottom of the reservoir 2820 is connected to a top of the valve2820. According to one embodiment, the valve 2820 can be a ball valve.The valve 2820 can have a single body design to minimize leak paths. Aheating element 2822 is applied to the valve 2820 to facilitateevacuation of waste from the reservoir 2802 above. In one embodiment,the heating element 2822 is in direct contact with the valve 2820. Forexample, in one embodiment, a cartridge heater 2822 is applied to thevalve 2820 to heat the valve 2820 by conduction.

In this embodiment, holes can be formed through the valve 2820 near theedges of the throat of the valve 2820 (as shown in the cut-away view ofthe valve 2820 in FIG. 35). The cartridge heater 2822, which can be anelongated pencil-like member, can be inserted into the holes and intothe valve 2820. Exemplary cartridge heaters 2822 that can be usedinclude cartridge heaters that are used for injection molding. Forexample, an exemplary cartridge heater 2822 is a FIREROD cartridgeheater having a diameter of about 0.125″. The cartridge heater 2822 isheated using electric current. Exemplary cartridge heater 2822 settingsinclude can be 48 VAC and 80 watts. Other voltage levels can be used ifdesired, such as 115 VAC. In the illustrated embodiment, the cartridgeheater 2822 includes two heating elements, one extending above thethroat of the valve 2820, the other extending below the throat. Aresistance temperature detector (RTD) monitor can be used to monitor theheat of the cartridge heater 2822. Persons skilled in the art willappreciate that other heating elements 2822, and manners of applyingheating elements 2822 to or into the valve 2820 can be utilized.

By directly contacting and heating the valve 2820, a cartridge heater2822 can heat waste promptly and act as a “hot plate” when the valve2820 is in the closed position. The valve 2820 is advantageously heatedso that the waste passing through the valve 2820 remains heated. This isparticularly advantageous for paraffin wax waste, which can begin tosolidify shortly after heat is not applied to wax. For example, paraffinwax can have a melting point of about 55°, and the heating element ofthe reservoir 2802 and/or the valve 2820 can set to heat the interior ofthe reservoir 2802 and/or valve 2820 to a temperature above this meltingpoint to maintain the solid paraffin wax and liquids or mixtures thereofin a semi-molten or molten state.

The bottom of the valve 2820 is connected to a waste exit chamber 2830.The waste exits the evacuation system 2800 via the chamber 2830 and intoa storage receptacle. The valve 2820 is connected to the chamber 2830via screws 2832 and a seal sleeve or liner 2834. The liner 2834 providesa seal between the valve 2820 and the waste exit chamber 2830. For thispurpose, the bottom of the valve 2820 can have a flexure to clamp to theliner 2834. The sealing material of the liner 2834 can be PFA(Perfluoroalkoxy). Alternatively, the liner 2834 material can be PTFETeflon. The screws 2832 prevent the liner 2834 from rotating ortranslating and to prevent the valve 2820 from translating. The screws2832 also make it easy to change the liner 2834 if the liner 2834 shouldneed replacing. A waste handle 2840 extends from the area below thevalve 2820 adjacent to the waste exit chamber 2830.

According to one embodiment, solid and liquid waste accumulates in thereservoir 2802 when the valve 2820 is closed. Vacuum is applied via port2806 to draw waste from the cell block engine 2803 until processing of acell block is complete, after which a chill or cooling cycle isinitiated. While the cell block is cooling, the waste can be heated,e.g., to 60° C. Prior to opening the valve 2820, the valve 2820 and/orvalve body can be heated, e.g., at 70° C. for about two minutes or othersuitable time. Different temperatures and heating durations can beutilized as necessary to achieve the desired level of heating.

A motor or other actuator 2840 is then activated to open the valve 2820.One exemplary motor 2840 includes a 3/16″ cross-pin/½″ drive shaft and amaximum torque of about 500 in-lbs (without pin yield). The motor 2840stall torque can be about 300 in-lbs. The valve 2820 can be actuatedwith about 50 in-lbs (max). The valve 2820 can be rotated by directdrive from the motor 2840. Exemplary motors 2840 include a 12 Volt, 1.2Amp, 300 in-lb Brush DC Gearmotor. Persons skilled in the art willappreciate that various motors and motor parameters and configurationscan be utilized with embodiments.

As a result of opening the valve 2820, the solid/liquid waste isevacuated from the waste collection area 2804, through the valve 2820,through the waste exit chamber 2830, and into collection receptacle orwaste tank positioned below the chamber 2830. For example, the valve2820 can have a throat having a diameter of about 0.8″ through whichwaste can pass. After evacuating the waste, it is not necessary tocontinue heating the valve 2820. Thus, the cartridge heater 2822 can bedeactivated to stop applying heat to the valve 2820. The valve 2820 isclosed, and the system 2800 is ready for preparation of the next cellblock, and the above heating and actuation process is repeated asnecessary.

Referring to FIGS. 35 and 39, to ensure that the valve 2820 is openedand closed properly, the rotational position of the valve 2820 can bemonitored via a limit switch or feedback system 2850. The output of thefeedback system 2850 can be used to control the motor 2840, which opensand closes the valve 2820. In one embodiment, a feedback system 2850 caninclude one or more sensors 3200. In the illustrated embodiment, thefeedback system 2850 includes two Hall Effect sensors 3200 and a wheel3202. The well 3204 includes a protrusion 3204 extending form the outeredge of the wheel 3202. A housing 3206 encloses and protects the sensors3200 from waste and other debris.

In the illustrated embodiment, the sensors 3200 are positioned at twelveo'clock and three o'clock positions, i.e., separated by 90 degrees. Inone embodiment, the valve 2820 is a ¼ turn valve, as shown in FIG. 35.Thus, at one position, the valve 2820 is closed, and rotating the valve2820 by 90 degrees opens the valve 2820. For purposes of explanation andillustration, the valve 2820 is closed when the wheel protrusion 3204 islocated at the twelve o'clock sensor 3200, and open when the protrusion3204 is located at the three o'clock sensor 3200.

In use, when the valve 2820 is closed, waste is accumulated in thereservoir 2802, and the motor 2840 can be activated to rotate the valve2820 to an open position. As the valve 2820 rotates, the wheel 3202connected thereto also moves from the closed twelve o'clock position toan open three o'clock position. Movement of the valve 2820 is limited bythe sensors 3200. More specifically, the protrusion 2304 of the wheel2302 moving 90 degrees from the twelve o'clock position to the threeo'clock position is detected by the second sensor 3200 at three o'clock.In this exemplary configuration, the protrusion 3204 interrupts a signaltransmitted between parts of an individual sensor 3200. Upon detectingthe protrusion 3204, the second sensor 3200 can generate a signal todeactivate the motor 2840 to limit rotation of the valve 2820 and keepthe valve open.

Similarly, when the waste has been evacuated, and the valve 2820 is tobe closed, the motor 2840 can be activated to rotate the valve 2820(which rotates the wheel 3202) back from the three o'clock position tothe twelve o'clock position. Movement of the valve 2820 in the oppositedirection is limited by the protrusion 3204 of the wheel 3202 moving 90degrees counterclockwise and being detected by the first sensor 3200 (asa result of an interrupted signal) at the twelve o'clock position. Upondetecting the protrusion 3204, the protrusion interrupts a signaltransmitted between parts of the individual sensor at the twelve o'clockposition, and in response, this sensor 3200 can generate a signal todeactivate the motor 2840. This limits rotation of the valve 2820 andkeeps the valve closed.

Persons skilled in the art will appreciate that different sensors 3200can be utilized to limit or control movement of the valve 2820. Further,persons skilled in the art will appreciate that different sensor 3200configurations can be utilized depending on the type of valve 2820 thatis utilized and the degree of rotation that is needed to open and closethe valve 2820. Further, the feedback system 2850 can be used to limitmovement of the valve 2820 between closed and open positions, or betweenclosed and partially open positions as necessary. Thus, the sensors3200, sensor 3200 configuration and valve 2820 shown in the figures areprovided for purposes of explanation and illustration, not limitation.

In a further alternative embodiment, FIG. 40 generally illustrates onealternative embodiment of a system 3300 in which solid waste (such asparaffin wax) and liquid waste (such as xylene and alcohol) areseparated and separately processed. In the illustrated embodiment, wastethat includes a mixture of solids (such as paraffin wax) and liquids(such as xylene and alcohol) can be divided or separated into solid andliquid components by a separator 3302, which outputs liquid waste 3304and solid waste 3306. Separated liquids 3304 can be stored in a liquidreservoir 2802 a, and separated solids can be stored in a solidsreservoir 2802 b.

Liquid waste is collected in the liquid reservoir 2802 a when the valve2820 a is closed. Similarly, solid waste is collected in the solidsreservoir 3312 when the valve 2820 b is closed. A foil heater 2812 canbe used to apply heat to the solids reservoir 2802 b, as described withreference to FIGS. 35-38 above. Heat can also be applied to the valve2820 b connected to the solids reservoir 2802 b via a cartridge heater2822, foil element, or other suitable heating element, as discussedabove. Solid waste can then be evacuated from the solids reservoir 2802b by opening the heated valve 2820 b. It may not be necessary to heatthe liquid reservoir 2802 a or the valve 2820 a positioned below theliquid reservoir 2802 a since the liquids from cell block processing donot solidify like wax. Thus, in the illustrated embodiment, only thesolids reservoir 2802 b and the valve 2820 b are heated for evacuatingsolid waste.

Further, persons skilled in the art will appreciate that in alternativeembodiments, the waste evacuation system shown in FIGS. 35-40 can bemodified. For example, although the above description refers to a ballvalve, or a rotating valve, other embodiments can use other types ofvalves, such as slide valves. Other alternative embodiments utilizedifferent heating elements. For example, although a valve was describedas being heated by a cartridge heater, in an alternative embodiment, avalve can also be heated by a foil heater. Similarly, although thereservoir was described as being heated by a foil heater. Thus, theheating elements used to heat the valve and the reservoir may be thesame or different heating elements. Additionally, although embodimentsare described in the context of cell block processing, embodiments maybe suitable for other applications, such as altering the flow rate of aviscous fluid by application of heat, and reducing friction of a thinfilm on a surface by application of heat.

The invention may be embodied in other specific forms besides and beyondthose described herein. The foregoing embodiments are therefore to beconsidered in all respects illustrative rather than limiting.

1. An automated method for forming a cell block in a cell blockcassette, the cassette having a collection well and a filter positionedacross a bottom surface of the collection well, the filter configured toretain cellular matter carried in a sample fluid flowing through thefilter, the method comprising: (a) dispensing a limited amount of samplefluid into the collection well; (b) determining a flow rate across thefilter based at least in part on a change in fluid level in thecollection well detected by a sensor; and (c) repeating steps (a) and(b) until a determined fluid flow rate across the filter indicates thata sufficient amount of cellular material has been retained by thefilter.
 2. The method of claim 1, further comprising applying a negativepressure differential across the filter to facilitate the sample fluidflow.
 3. The method of claim 1, further comprising selectively passingair through the filter in a reverse direction to the sample fluid flowto thereby push at least some retained cellular mater off of a topsurface of the filter, at least in part in response to a determinedfluid flow rate across the filter.
 4. The method of claim 1, wherein theamount of sample fluid dispensed during a repeated step (a) is based atleast in part on one or both of an existing fluid level in the fluidwell and a prior determined flow rate across the filter.
 5. The methodof claim 1, wherein the sample fluid is dispensed into the collectionwell by an automated arm assembly.
 6. The method of claim 5, theautomated arm assembly comprising a pipette tip holder configured toselectively retrieve, carry and dispose of pipette tips, the automatedarm assembly configured to (i) retrieve a pipette tip, (ii) position theretrieved pipette tip to draw fluid from a sample vial, and (iii)dispense the drawn sample fluid into the collection well.
 7. The methodof claim 1, further comprising, after a sufficient amount of cellularmaterial has been retained by the filter, dispensing one or more liquidreagents into the collection well to treat the cellular materialretained by the filter; dispensing liquefied paraffin into thecollection well to embed the treated cellular material; and allowing theparaffin to solidify in the collection well to embed the cellularmaterial in solid paraffin.
 8. The method of claim 7, wherein allowingthe paraffin to solidify comprises chilling the collection well tothereby chill the paraffin.
 9. The method of claim 7, further comprisingchilling the already solidified paraffin to cause thermal contraction ofsame; and upon chilling the solidified paraffin, separating the filterfrom the paraffin.
 10. The method of claim 7, further comprising,removing filter from the paraffin-embedded cellular material; placingthe paraffin-embedded cellular material atop an additional amount ofparaffin; and controllably heating to thereby soften and at leastpartially blend together the embedding paraffin and additional paraffin,without softening or liquefying the embedding paraffin to a point thatthe retained cellular material therein breaks apart and disbursesthrough the embedding paraffin.
 11. The method of claim 10, furthercomprising controllably cooling to thereby bond the additional paraffinto the embedding paraffin.
 13. The method of claim 7, wherein the filteris positioned beyond a bottom surface of the main cassette body so thatthe paraffin-embedded cellular matter is also located beyond the maincassette bottom surface.
 14. The method of claim 7, wherein prior todispensing liquefied paraffin into the collection well, a thermallyconductive substrate underlying the filter is heated to thereby heat thecontents of the collection well.
 15. An automated method for forming acell block in a cell block cassette, the cassette having a collectionwell and a filter positioned across a bottom surface of the collectionwell, the filter configured to retain cellular matter carried in asample fluid flowing through the filter, the method comprising:dispensing sample fluid into the collection well; dispensing one or moreliquid reagents into the collection well to treat cellular materialretained from the sample fluid by the filter; heating a thermallyconductive substrate underlying the filter to thereby heat the contentsof the collection well; after heating the contents of the collectionwell, dispensing liquefied paraffin into the collection well to embedthe treated cellular material; and after dispensing liquefied paraffininto the collection well to embed the treated cellular material,chilling the contents of the collection well to solidify the paraffin.16. The method of claim 15, further comprising chilling the alreadysolidified paraffin to cause thermal contraction of same; and uponchilling the solidified paraffin, separating the filter from theparaffin.
 17. The method of claim 15, further comprising, removingfilter from the paraffin-embedded cellular material; placing theparaffin-embedded cellular material atop an additional amount ofparaffin; and controllably heating to thereby soften and at leastpartially blend together the embedding paraffin and additional paraffin,without softening or liquefying the embedding paraffin to a point thatthe retained cellular material therein breaks apart and disbursesthrough the embedding paraffin.
 18. The method of claim 17, furthercomprising controllably cooling to thereby bond the additional paraffinto the embedding paraffin.
 19. An automated method for forming a cellblock in a cell block cassette, the cassette having a collection welland a filter positioned across a bottom surface of the collection well,the filter configured to retain cellular matter carried in a samplefluid flowing through the filter, the method comprising: (a) using anautomated arm assembly to dispense a limited amount of sample fluid intothe collection well; (b) applying a negative pressure differentialacross the filter to cause the sample fluid flow across the filter; (c)determining a flow rate across the filter based at least in part on achange in fluid level in the collection well detected by a sensor; and(d) repeating steps (a)-(c) until a determined fluid flow rate acrossthe filter indicates that a sufficient amount of cellular material hasbeen retained by the filter.
 20. The method of claim 19, furthercomprising selectively passing air through the filter in a reversedirection to the sample fluid flow to thereby push at least someretained cellular mater off of a top surface of the filter, at least inpart in response to a determined fluid flow rate across the filter. 21.The method of claim 19, wherein the amount of sample fluid dispensedduring a repeated step (a) is based at least in part on one or both ofan existing fluid level in the fluid well and a prior determined flowrate across the filter.
 22. The method of claim 19, the automated armassembly comprising a pipette tip holder configured to selectivelyretrieve, carry and dispose of pipette tips, the automated arm assemblyconfigured to (i) retrieve a pipette tip, (ii) position the retrievedpipette tip to draw fluid from a sample vial, and (iii) dispense thedrawn sample fluid into the collection well.
 23. The method of claim 19,further comprising, after a sufficient amount of cellular material hasbeen retained by the filter, dispensing one or more liquid reagents intothe collection well to treat the cellular material retained by thefilter; dispensing liquefied paraffin into the collection well to embedthe treated cellular material; allowing the paraffin to solidify in thecollection well to embed the cellular material in solid paraffin; andmanually separating the filter from the solidified paraffin.
 24. Themethod of claim 23, wherein the step of manually removing the filterfrom the solidified paraffin comprises chilling the solidified paraffinto cause thermal contraction of same.
 25. The method of claim 23,further comprising, placing the paraffin-embedded cellular material atopan additional amount of paraffin; controllably heating to thereby softenand at least partially blend together the embedding paraffin andadditional paraffin, without softening or liquefying the embeddingparaffin to a point that the retained cellular material therein breaksapart and disburses through the embedding paraffin; and controllablycooling to thereby bond the additional paraffin to the embeddingparaffin.